The present invention relates to compositions and methods for the inhibition of cellular proliferation. More particularly, the present invention relates to the use of ligands that bind to components of the blood coagulation/clotting pathway, and inhibit angiogenesis and angiogenesis-related diseases.
Cellular proliferation is a normal ongoing process in all living organisms and is one that involves numerous factors and signals that are delicately balanced to maintain regular cellular cycles. The general process of cell division is one that consists of two sequential processes: nuclear division (mitosis), and cytoplasmic division (cytokinesis). Because organisms are continually growing and replacing cells, cellular proliferation is a central process that is vital to the normal functioning of almost all biological processes. Whether or not mammalian cells will grow and divide is determined by a variety of feedback control mechanisms, which include the availability of space in which a cell can grow, and the secretion of specific stimulatory and inhibitory factors in the immediate environment.
When normal cellular proliferation is disturbed or somehow disrupted, the results can affect an array of biological functions. Disruption of proliferation could be due to a myriad of factors such as the absence or overabundance of various signaling chemicals or presence of altered environments. Some disorders characterized by abnormal cellular proliferation include cancer, abnormal development of embryos, improper formation of the corpus luteum, difficulty in wound healing as well as malfunctioning of inflammatory and immune responses.
Cancer is characterized by abnormal cellular proliferation. Cancer cells exhibit a number of properties that make them dangerous to the host, often including an ability to invade other tissues and to induce capillary ingrowth, which assures that the proliferating cancer cells have an adequate supply of blood. One of the defining features of cancer cells is that they respond abnormally to control mechanisms that regulate the division of normal cells and continue to divide in a relatively uncontrolled fashion until they kill the host.
Angiogenesis and angiogenesis-related diseases are closely affected by cellular proliferation. As used herein, the term xe2x80x9cangiogenesisxe2x80x9d means the generation of new blood vessels into a tissue or organ. Under normal physiological conditions, humans or animals undergo angiogenesis only in very specific restricted situations. For example, angiogenesis is normally observed in wound healing, fetal and embryonal development and formation of the corpus luteum, endometrium and placenta. The term xe2x80x9cendotheliumxe2x80x9d is defined herein as a thin layer of flat cells that lines serous cavities, lymph vessels, and blood vessels. These cells are defined herein as xe2x80x9cendothelial cellsxe2x80x9d. The term xe2x80x9cendothelial inhibiting activityxe2x80x9d means the capability of a molecule to inhibit angiogenesis in general. The inhibition of endothelial cell proliferation also results in an inhibition of angiogenesis.
Both controlled and uncontrolled angiogenesis are thought to proceed in a similar manner. Endothelial cells and pericytes, surrounded by a basement membrane, form capillary blood vessels. Angiogenesis begins with the erosion of the basement membrane by enzymes released by endothelial cells and leukocytes. The endothelial cells, which line the lumen of blood vessels, then protrude through the basement membrane. Angiogenic stimulants induce the endothelial cells to migrate through the eroded basement membrane. The migrating cells form a xe2x80x9csproutxe2x80x9d off the parent blood vessel, where the endothelial cells undergo mitosis and proliferate. The endothelial sprouts merge with each other to form capillary loops, creating the new blood vessel.
Persistent, unregulated angiogenesis occurs in a multiplicity of disease states, tumor metastasis and abnormal growth by endothelial cells and supports the pathological damage seen in these conditions. The diverse pathological disease states in which unregulated angiogenesis is present have been grouped together as angiogenic-dependent, angiogenic-associated, or angiogenic-related diseases. These diseases are a result of abnormal or undesirable cell proliferation, particularly endothelial cell proliferation.
The hypothesis that tumor growth is angiogenesis-dependent was first proposed in 1971 by Judah Folkman (N. Engl. Jour. Med. 285:1182 1186, 1971). In its simplest terms the hypothesis proposes that once tumor xe2x80x9ctakexe2x80x9d has occurred, every increase in tumor cell population must be preceded by an increase in new capillaries converging on the tumor. Tumor xe2x80x9ctakexe2x80x9d is currently understood to indicate a prevascular phase of tumor growth in which a population of tumor cells occupying a few cubic millimeters volume and not exceeding a few million cells, survives on existing host microvessels. Expansion of tumor volume beyond this phase requires the induction of new capillary blood vessels. For example, pulmonary micrometastases in the early prevascular phase in mice would be undetectable except by high power microscopy on histological sections. Further indirect evidence supporting the concept that tumor growth is angiogenesis dependent is found in U.S. Pat. Nos. 5,639,725, 5,629,327, 5,792,845, 5,733,876, and 5,854,205, all of which are incorporated herein by reference.
Thus, it is clear that cellular proliferation, particularly endothelial cell proliferation, and most particularly angiogenesis, plays a major role in the metastasis of a cancer. If this abnormal or undesirable proliferation activity could be repressed, inhibited, or eliminated, then the tumor, although present, would not grow. In the disease state, prevention of abnormal or undesirable cellular proliferation and angiogenesis could avert the damage caused by the invasion of the new microvascular system. Therapies directed at control of the cellular proliferative processes could lead to the abrogation or mitigation of these diseases.
What is needed are compositions and methods which can inhibit abnormal or undesirable cellular proliferation, especially the growth of blood vessels into tumors. The compositions should be able to overcome the activity of endogenous growth factors in premetastatic tumors and prevent the formation of the capillaries in the tumors thereby inhibiting the development of disease and the growth of tumors. The compositions should also be able to modulate the formation of capillaries in angiogenic processes, such as wound healing and reproduction. Finally, the compositions and methods for inhibiting cellular proliferation should preferably be non-toxic and produce few side effects.
Compositions and methods are provided that are effective in inhibiting abnormal or undesirable cell proliferation, particularly endothelial cell proliferation and angiogenesis related to neovascularization and tumor growth. The compositions comprise naturally occurring or synthetic proteins, peptides, or protein fragments containing all, or active fragments of ligands that bind components of the blood coagulation/clotting pathway, optionally provided in a pharmaceutically acceptable carrier.
Representative ligands useful for the present invention comprise proteins or peptides that bind components of both the intrinsic and extrinsic blood clotting pathways such tissue factor (TF), and/or TF cofactor complexes, factor V, factor VIII, factor XII, factor XI, factor X, factor IX, factor VIIa, thrombin, fibrinogen and fibrin.
Preferred ligand compositions of the present invention, include but are not limited to, proteins comprising Kunitz domain proteins, non-Kunitz domain proteins, Kringle-rich proteins, TF cofactors (i.e. factor VIIa, phospholipids, gangliosides), TF antagonists (i.e. antibodies), and any other molecules that bind TF. More particularly, the compositions of the present invention comprise TFPI, protein S, protein Z, protein Z inhibitor, protein C, activated protein C, protein C inhibitor, prothrombin, group II secretory phospholipase A2, complement protein C4b, protease nexin-1, beta2-glycoprotein I, and serpins anticoagulants such as antithrombin and heparin cofactor II.
Preferably, the protein, peptide or protein fragment contains all or an active portion of the above identified proteins. The term xe2x80x9cactive fragmentxe2x80x9d, as used herein, means a portion of a protein that inhibits abnormal or undesirable cell proliferation, more specifically inhibits endothelial cell proliferation. Also included in the present invention are homologs, peptides, or protein fragments, or combinations thereof of the above-identified proteins, that inhibit abnormal or undesirable cell proliferation. Most preferably, the protein or peptide comprises TF or factor Xa binding ligand, or an active fragment thereof.
Though not wishing to be bound by the following theory, it is believed that by inhibiting endothelial cell proliferation, the methods and compositions described herein are useful for inhibiting tumor growth and metastasis by blocking tumor vascularization. The methods provided herein for treating diseases and processes mediated by undesired and uncontrolled cell proliferation, such as cancer, involve administering to a human or animal the composition described herein in a dosage sufficient to inhibit cell proliferation, particularly endothelial cell proliferation. The methods are especially useful for treating or repressing the growth of tumors, particularly by inhibiting angiogenesis. Administration of the compositions to a human or animal having prevascularized metastasized tumors is useful for preventing the growth or expansion of such tumors.
Accordingly, it is an object of the present invention to provide methods and compositions for treating diseases and processes that are mediated by abnormal or undesirable cellular proliferation.
It is another object of the present invention to provide methods and compositions for treating or repressing the growth of a cancer.
It is yet another object of the present invention to provide methods and compositions for therapy of cancer that has minimal side effects.
It is another object of the present invention to provide methods and compositions for treating diseases and processes that are mediated by angiogenesis.
Yet another object of the present invention is to provide methods and compositions comprising the use of proteins, peptides, active fragments and homologs thereof that inhibit endothelial cell proliferation.
Another object of the present invention is to provide methods and compositions for treating diseases and processes that are mediated by angiogenesis by administrating antiangiogenic compounds comprising ligands that bind components of the blood coagulation/clotting pathway.
It is another object of the present invention to provide methods and compositions for treating diseases and processes that are mediated by angiogenesis by administrating antiangiogenic compounds comprising ligands that bind components of the intrinsic and/or extrinsic blood clotting pathways wherein in such components comprise tissue factor (TF), and/or TF cofactor complexes, factor V, factor VIII, factor XII, factor XI, factor X, factor IX, factor VIIa, thrombin, fibrinogen and fibrin.
It is a further object of the present invention to provide methods and compositions for treating diseases and processes that are mediated by angiogenesis by administrating antiangiogenic compounds comprising ligands, wherein the ligands comprise Kunitz domain proteins, non-Kunitz domain proteins, Kringle-rich proteins, TF cofactors, TF antagonists, factor VIIa antagonists or inhibitors and factor Xa antagonists or inhibitors.
It is another object of the present invention to provide methods and compositions for treating diseases and processes that are mediated by angiogenesis comprising administration of antiangiogenic compounds comprising TFPI, protein S, protein Z, protein Z inhibitor, protein C, activated protein C, protein C inhibitor, prothrombin, group II secretory phospholipase A2, complement protein C4b, protease nexin-1, beta2-glycoprotein I, and serpins anticoagulants (such as antithrombin and heparin cofactor II) and inhibitors of factors TF, TF/VIIa, VIIa, Xa.
It is still another object of the present invention to provide antiangiogenic compositions comprising ligands that bind components of the intrinsic and extrinsic blood clotting pathways wherein the compositions further comprise pharmaceutically acceptable carriers.
Yet another object of the present invention is to provide antiangiogenic compositions comprising ligands that bind components of the intrinsic and/or extrinsic blood clotting pathways wherein the compositions further comprise pharmaceutically acceptable carriers that may be administered intramuscularly, intravenously, transdermally, orally, or subcutaneously.
It is yet another object of the present invention to provide compositions and methods for treating diseases and processes that are mediated by angiogenesis including, but not limited to, hemangioma, solid tumors, blood borne tumors, leukemia, metastasis, telangiectasia, psoriasis, scleroderma, pyogenic granuloma, myocardial angiogenesis, Crohn""s disease, plaque neovascularization, arteriovenous malformations, corneal diseases, rubeosis, neovascular glaucoma, diabetic retinopathy, retrolental fibroplasia, arthritis, diabetic neovascularization, macular degeneration, wound healing, peptic ulcer, Helicobacter related diseases, fractures, keloids, vasculogenesis, hematopoiesis, ovulation, menstruation, placentation, and cat scratch fever.
These and other objects, features and advantages of the present invention will become apparent after a review of the following detailed description of the disclosed embodiment and the appended claims.